Obesity is a major health problem in many countries, leading to serious comorbidities. The gut is the first point in the body at which the quantity and quality of ingested nutrients is monitored. Therefore, it is important to understand the mechanisms by which nutrients are sensed in the gut, the neural pathways activated, and how these may be altered by long term changes in diet leading to increased food intake and obesity. Ingestion of high fat diets and increased adiposity is associated with an inflammatory state and diet-induced obesity is associated with low chronic plasma level of lipopolysaccharide (LPS), a breakdown product of the cell wall of Gram-negative bacteria. This observation, together with recent evidence showing a change in the gut microbiota in obesity and following ingestion of high fat diets, suggests a role for LPS in the development or maintenance of obesity. Vagal afferents express TLR4 (15), but how LPS influences CCK-dependent activation of the vagal afferent pathway or expression of orexigenic and anorexigenic receptors has not been explored. The goal of the proposal is to determine the mechanism by which activation of the vagal afferent pathway by CCK and lipid is attenuated by long term ingestion of a high fat diet, leading to a reduction in short term satiety, hyperphagia and altered body weight regulation. The hypothesis to be tested is that long term ingestion of a high fat diet inhibits the CCK-induced changes in receptor and peptide expression in vagal afferent neurons, resulting in the persistence of an orexigenic phenotype and reduced satiety. The first specific aim will address the role of LPS and TLR4 in the reduced sensitivity of the vagal afferent pathway to CCK in high fat diet-induced obesity. Experiments will determine the effect of chronic ingestion of a high fat diet on intestinal function that lead to an increase in plasma levels of LPS and on change in the phenotype of vagal afferent neurons and whether chronic administration of LPS recapitulates the diet-induced obese phenotype and attenuates CCK-induced changes in vagal afferent function. The role of TLR4 in reduced sensitivity of the vagal afferent pathway to CCK to HF diets will be determined using TLR4 null mice. The second specific aim will address changes at the level of the vagal afferent neurons to determine the mechanism by which a high fat diet inhibits the CCK- or feeding-induced change switch from an orexigenic to an anorexigenic phenotype in vagal afferent neurons. Experiments will determine that the switch from orexigenic to anorexigenic phenotype induced by CCK or feeding in vagal afferent neurons is abolished or attenuated in HF diet-induced obesity and that this occurs either leptin resistance or an upregulation of the ghrelin receptor in vagal afferent neurons, or a decrease production of intestinal apolipoprotein A-IV. The significance of this proposal is that a greater understanding of the plasticity of the vagal afferent pathway, together with the possibility of targeting a luminal site of action of an obesity treatment, will provide better targets for the treatment of obesity and other disorders that involve the sensory innervation of the gut, such as functional bowel disease.